CN104186957A - Method for improving forage nutritional value of cottonseed meal through microbial fermentation - Google Patents
Method for improving forage nutritional value of cottonseed meal through microbial fermentation Download PDFInfo
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- CN104186957A CN104186957A CN201410518197.8A CN201410518197A CN104186957A CN 104186957 A CN104186957 A CN 104186957A CN 201410518197 A CN201410518197 A CN 201410518197A CN 104186957 A CN104186957 A CN 104186957A
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention utilizes a microbial fermentation technology to improve the forage nutritional value of the cottonseed meal, so as to improve the forage quality of the cottonseed meal, increase the addition proportion in the forage and realize the promotion of the protein nutritional value and the feeding value of the cottonseed meal, aiming at the disadvantages that cottonseed meal is rich in antinutritional factors of free gossypol, cellulose, oligosaccharide, phytic acid and the like, the protein quality is relatively poor and the content of essential amino acids of methionine and the like is low and unbalanced, so that the large-scale forage of cottonseed meal is restricted. The method for improving forage nutritional value of cottonseed meal through the microbial fermentation adopts the technology of 'composite enzymatic hydrolysis and multi-strain three-step solid-state fermentation', so as to prepare the fermented cottonseed meal product that is high in the free gossypol, cellulose degradation degree and the protein content; the cottonseed meal product is rich in nutritional ingredients of probiotics, vitamins, enzyme, amino acid, short peptide, high-grade protein and the like; the method for improving the forage nutritional value of cottonseed meal through the microbial fermentation has the advantages of prompting growth and development, improving the utilization ratio of the forage, enhancing immunization, improving intestinal micro ecology, preventing diseases and the like. The method for improving forage nutritional value of cottonseed meal through the microbial fermentation adopts the solid-state fermentation; the manufacturing technology is simple; the energy consumption is low; zero environmental pollution is generated; and the investment is small. Therefore, mass production is easy.
Description
Technical field
The present invention relates to the method that microorganism fermentation improves cotton dregs Forage Nutritive Value, belong to field of feed processing.
Background technology
Protein raw material is the basis of feed industrial development, the surging bottleneck that has become restriction animal husbandry development of in short supply and price of China's protein raw material.Current raising and the attention to quality of life along with people's living standard, demand to meat, egg, milk, aquatic products is increasing, grain and production of fodder face a severe test, feed protein resource will be day by day deficient, therefore improve grain to the utilization rate of transformation efficiency and the feed of livestock products, expand feed resource, develop unconventional feed extremely urgent.And the Critical policies alleviating the problems is exactly for existing protein sources, improve its utilization rate, excavate as much as possible the nutritional labeling utilized of feed, fundamentally alleviate the situation that feedstuff is in short supply.
China is Yi Gechanmian big country, and nearly 1,000 ten thousand tons of cottonseed, more than 600 ten thousand tons of cotton dregs are produced in every annual.In cotton dregs, crude protein content is about 44%, and consumption figure accounts for 8% of protein feed, is important protein feed resource.But cotton dregs, containing noxious material gossypol, can produce obvious harmful effect to growth of animal, development and fecundity etc.; And contain the ANFs such as cellulose, compound sugar (gossypose, stachyose), phytic acid; Crude protein content is high, but protein quality is poor, methionine etc., essential amino acids content is low and uneven, above-mentioned effects limit the extensive uses of cotton dregs in animal husbandry.All the time, in order to improve the application of cotton dregs in feed, researcher has carried out the research work of the screening of gossypol high-efficiency detoxicating bacterial strain and detoxification efficiency thereof etc., but cotton dregs nutritive value is promoted and lacks significant effect.Efficient Development utilizes cotton dregs, realizes the lifting of cottonseed meal nutritive value and feeding value, and the development of agricultural byproducts processing industry and animal husbandry is had to very important facilitation, significant.
Summary of the invention
The present invention is directed to and in cotton dregs, be rich in the ANFs such as free gossypol, cellulose, compound sugar, phytic acid; and protein quality is poor; the essential amino acids content such as methionine are low and uneven; restricted the feeding drawback of cotton dregs scale; utilize microbial fermentation technology; according to the accumulation of the useful metabolites such as the variation of cotton dregs nutritional labeling and essential amino acid, lactic acid; improve the Forage Nutritive Value of cotton dregs; thereby improve the feeding quality of cotton dregs and the adding proportion in feed, realize the lifting of cottonseed meal nutritive value and feeding value.
Above-mentioned purpose of the present invention is achieved by the following technical programs:
The invention provides the method that microorganism fermentation improves cotton dregs Forage Nutritive Value, it is characterized in that specifically comprising the following steps:
(1) pretreatment: choose without the cotton dregs that go mouldy, pulverized 40 mesh sieves, adjust cotton dregs water content 12～18% left and right;
(2) enzymolysis: adjust cotton dregs pH4.0～5.0, add cellulase, zytase, enzyme concentration (mass ratio) is 2～3%, 1～2%, mixes, enzymolysis 16～24h, and hydrolysis temperature is controlled at 40～50 ℃;
(3) preparation solid medium: add 3～5%(with respect to cotton dregs mass ratio in the cotton dregs after enzymolysis, molasses, 2～3% corn steep liquors, 1.5～2% urea, 1～1.5% ammonium sulfate, 0.1～0.2% potassium dihydrogen phosphate down together), adjust moisture content in medium 30～40%, adjusting initial pH is 7.0 left and right;
(4) aspergillus activation: get fresh wheat bran, with 60 mesh sieves, sieve and remove fine powder, in wheat bran: water=1:1.0～1.3 ratio adds water, mix thoroughly to without dry powder again without clustering phenomena.After mixing thoroughly, be distributed in 2000mL triangular flask, every about 100g of the bottled wet wheat bran of 2000mL triangle, sterilizing 40～60min under 0.1MPa gauge pressure, be cooled to 35 ℃, in each triangular flask, access inclined-plane aspergillus oryzae cv-8, aspergillus niger sp-1 spore that 1-2 ring has activated, the temperature control of culture medium product is in (35 ± 1) ℃. every 12-24h shaking flask once, after growing, spore stops preparing spore suspension;
(5) solid fermentations: aspergillus oryzae, aspergillus niger are inoculated in mix in 2～3%, 3～5% ratio, mix 35～40 ℃ of cultivations thoroughly, aerlbic culture 3～5 days;
(6) candida utili, saccharomyces cerevisiae activation: candida utili, saccharomyces cerevisiae are received respectively in YPD culture medium, and 30 ℃, under 180r/min rotating speed, constant temperature shaking flask is 24 hours; Then with 1:100 ratio, seed liquor is received in YPD culture medium, obtained saccharomyces cerevisiae and candida utili liquid seeds;
(7) bacillus subtilis activation: adopt LB culture medium to cultivate 24h under 28～32 ℃ of conditions, make liquid seeds;
(8) secondary solid fermentation: candida utili, saccharomyces cerevisiae, bacillus subtilis are seeded to culture medium by 2～3%, 1～2%, 2～3% inoculative proportions, cultivate 2～3 days for about 28 ℃;
(9) three solid fermentations: solid lactic acid bacterium powder is seeded to culture medium by 1～2% inoculative proportion, and 26～30 ℃ of anaerobism are cultivated 24h left and right;
(10) dry: after fermentation ends, low-temperature aeration is dry;
(11) pulverize: dried fermentation cotton dregs are pulverized and got product.
In above-mentioned steps (3), solid medium is without sterilizing.
The invention provides the method that microorganism fermentation improves cotton dregs Forage Nutritive Value, promoted greatly the Forage Nutritive Value of cotton dregs: in fermentation cotton dregs, crude protein content rises to 52.2% after fermentation by 43.6% before fermenting; Utilize automatic amino acid analyzer to measure the variation that in the cotton dregs of fermentation front and back, amino acid forms, total amino acid content rises to 50.70% after fermentation by 37.79% before fermenting, wherein TEAA rises to 15.31% after fermentation by 11.93% before fermenting, wherein lysine content rises to 2.28% after fermentation by 1.78% before fermenting, methionine rises to 0.48% after fermentation by 0.31% before fermenting, isoleucine rises to 1.82% after fermentation by 1.41% before fermenting, and arginine rises to fermentation rear 6.46% by 5.10% before fermenting; Measure fermented cotton cake protein matter and form, little peptide content rises to 21.28% after fermentation by 3.46% before fermenting; In fermentation cotton dregs, lactic acid content reaches 7.0%; In addition, in fermentation cotton dregs, contain the nutritional labelings such as a large amount of useful viable bacterias, multivitamin, enzyme.
The invention provides the method that microorganism fermentation improves cotton dregs Forage Nutritive Value, the ANFs such as cotton dregs Free Gossypol, cellulose, xylan, phytic acid of effectively having degraded: free gossypol drops to the 16mg/Kg after fermentation by the 520mg/Kg before fermenting, and virus elimination rate reaches 96.92%; Gossypose content is reduced to degradable after fermentation by 3.6% before fermenting; Crude fibre is reduced to 2.5% after fermentation by 12.0% before fermenting; Phytic acid content is reduced to 0.18% after fermentation by 1.85% before fermenting, and degradation rate reaches 90.27%.
The invention provides the method that microorganism fermentation improves cotton dregs Forage Nutritive Value, its advantage is: process by cotton dregs complex enzyme hydrolysis (1), the difficult cellulose utilizing of microorganism, xylan etc. are degraded to reduced sugar, and content of reducing sugar rises to microbial growth breeding sufficient carbon source is provided; (2) aspergillus niger and aspergillus oryzae are the most significant bacterial strains of current known degrading straw effect, the mutualistic symbiosis relation of mould and saccharomycete Mixed culture improves a lot cottonseed meal content, saccharomycete not only can play derepression, and the growth of fungi is just had and imitated, analyze the mould assimilation starches such as reason aspergillus oryzae, aspergillus niger and cellulosic ability strong, the structural carbohydrate of cotton dregs can be degraded to the monosaccharide materials such as the available monosaccharide and disaccharide of yeast, make yeast be able to good growth; In addition,, in mixed fermentation with various bacterium, the sugar that enzymatic catalysis generates is fermented immediately sugared Institute of Micro-biology and utilizes, so just maintained the concentration of degradation product, eliminate the effect of checking of the degradation product that enzymatic synthesis effect is subject to, also removed the feedback inhibition of reacting final product to enzyme, shortened sweat; (3) adopt many bacterium mixed fungus fermentation, combination strain fermentation has increased the many gene functions in fermentation, by the combination of different metabolic ability, complete the complicated metabolism that single bacterial classification has been difficult to, by the reciprocal and laterality life between microorganism, enhance productivity, the existence of saccharomyces cerevisiae and candida utili has promoted the degradation of mould, and the amount reproduction of yeast has increased the mycoprotein in product, bacillus subtilis has been improved probio content in product, has improved the effect of product; Lactic acid bacteria generation lactic acid etc. can improve the palatability of fermentation cotton dregs.Cooperative fermentation form improves and has played important positive role the quality of effective conversion of various raw materials, protein feed; (4) adopt stepwise fermentation, fermentation strain requires different to temperature, pH, logical oxygen etc., and different reaction controlled condition has improved the growth rate of thalline, has shortened fermentation time, has promoted ferment effect; (5) in sweat without sterilizing, thereby the problem of having avoided the amino acid that causes because of high temperature in sterilization process and carbohydrate reaction to cause protein digestibility to decline; (6) in product, be rich in the nutritional labelings such as probio, vitamin, enzyme and amino acid, small peptide, good protein, there is enhancing development, improve efficiency of feed utilization, strengthen immunity, improve intestinal microecology, prevent disease, reduce the advantages such as environmental pollution; (7) adopt solid state fermentation, production technology is simple, and energy consumption is low, non-environmental-pollution, investment are little, be easy to large-scale production.
The specific embodiment
embodiment 1:
Microorganism fermentation improves the method for cotton dregs Forage Nutritive Value, and concrete preparation process is as follows:
(1) pretreatment: choose without the cotton dregs that go mouldy, pulverized 40 mesh sieves, adjust cotton dregs water content 12～18% left and right;
(2) enzymolysis: adjust cotton dregs pH4.0～5.0, add cellulase, zytase, enzyme concentration (mass ratio) is 3%, 1%, mixes, enzymolysis 16h, and hydrolysis temperature is controlled at 40～50 ℃;
(3) preparation solid medium: add 5%(with respect to cotton dregs mass ratio in the cotton dregs after enzymolysis, molasses, 2% corn steep liquor, 2% urea, 1% ammonium sulfate, 0.2% potassium dihydrogen phosphate down together), adjust moisture content in medium 30～40%, adjusting initial pH is 7.0 left and right;
(4) aspergillus activation: get fresh wheat bran, with 60 mesh sieves, sieve and remove fine powder, in wheat bran: water=1:1.0 ratio adds water, mix thoroughly to without dry powder again without clustering phenomena.After mixing thoroughly, be distributed in 2000mL triangular flask, every about 100g of the bottled wet wheat bran of 2000mL triangle, sterilizing 40～60min under 0.1MPa gauge pressure, be cooled to 35 ℃, in each triangular flask, access inclined-plane aspergillus oryzae cv-8, aspergillus niger sp-1 spore that 1-2 ring has activated, the temperature control of culture medium product is in (35 ± 1) ℃. every 12-24h shaking flask once, after growing, spore stops preparing spore suspension;
(5) solid fermentations: aspergillus oryzae, aspergillus niger are inoculated in mix in 3%, 3% ratio, mix 35～40 ℃ of cultivations thoroughly, aerlbic culture 3～5 days;
(6) candida utili, saccharomyces cerevisiae activation: candida utili, saccharomyces cerevisiae are received respectively in YPD culture medium, and 30 ℃, under 180r/min rotating speed, constant temperature shaking flask is 24 hours; Then with 1:100 ratio, seed liquor is received in YPD culture medium, obtained saccharomyces cerevisiae and candida utili liquid seeds;
(7) bacillus subtilis activation: adopt LB culture medium to cultivate 24h under 28～32 ℃ of conditions, make liquid seeds;
(8) secondary solid fermentation: candida utili, saccharomyces cerevisiae, bacillus subtilis are seeded to culture medium by 3%, 1%, 3% inoculative proportion, cultivate 2～3 days for about 28 ℃;
(9) three solid fermentations: solid lactic acid bacterium powder is seeded to culture medium by 2% inoculative proportion, and 26～30 ℃ of anaerobism are cultivated 24h left and right;
(10) dry: after fermentation ends, low-temperature aeration is dry;
(11) pulverize: dried fermentation cotton dregs are pulverized and got product.
embodiment 2:
(1) pretreatment: choose without the cotton dregs that go mouldy, pulverized 40 mesh sieves, adjust cotton dregs water content 12～18% left and right;
(2) enzymolysis: adjust cotton dregs pH4.0～5.0, add cellulase, zytase, enzyme concentration (mass ratio) is 2%, 2%, mixes, enzymolysis 16～24h, and hydrolysis temperature is controlled at 40～50 ℃;
(3) preparation solid medium: add 3%(with respect to cotton dregs mass ratio in the cotton dregs after enzymolysis, molasses, 3% corn steep liquor, 1.5% urea, 1.5% ammonium sulfate, 0.1% potassium dihydrogen phosphate down together), adjust moisture content in medium 30～40%, adjusting initial pH is 7.0 left and right;
(4) aspergillus activation: get fresh wheat bran, with 60 mesh sieves, sieve and remove fine powder, in wheat bran: water=1:1.3 ratio adds water, mix thoroughly to without dry powder again without clustering phenomena.After mixing thoroughly, be distributed in 2000mL triangular flask, every about 100g of the bottled wet wheat bran of 2000mL triangle, sterilizing 40～60min under 0.1MPa gauge pressure, be cooled to 35 ℃, in each triangular flask, access inclined-plane aspergillus oryzae cv-8, aspergillus niger sp-1 spore that 1-2 ring has activated, the temperature control of culture medium product is in (35 ± 1) ℃. every 12-24h shaking flask once, after growing, spore stops preparing spore suspension;
(5) solid fermentations: aspergillus oryzae, aspergillus niger are inoculated in mix in 2%, 5% ratio, mix 35～40 ℃ of cultivations thoroughly, aerlbic culture 3～5 days;
(6) candida utili, saccharomyces cerevisiae activation: candida utili, saccharomyces cerevisiae are received respectively in YPD culture medium, and 30 ℃, under 180r/min rotating speed, constant temperature shaking flask is 24 hours; Then with 1:100 ratio, seed liquor is received in YPD culture medium, obtained saccharomyces cerevisiae and candida utili liquid seeds;
(7) bacillus subtilis activation: adopt LB culture medium to cultivate 24h under 28～32 ℃ of conditions, make liquid seeds;
(8) secondary solid fermentation: candida utili, saccharomyces cerevisiae, bacillus subtilis are seeded to culture medium by 2%, 2%, 2% inoculative proportion, cultivate 2～3 days for about 28 ℃;
(9) three solid fermentations: solid lactic acid bacterium powder is seeded to culture medium by 1% inoculative proportion, and 26～30 ℃ of anaerobism are cultivated 24h left and right;
(10) dry: after fermentation ends, low-temperature aeration is dry;
(11) pulverize: dried fermentation cotton dregs are pulverized and got product.
embodiment 3:
(1) pretreatment: choose without the cotton dregs that go mouldy, pulverized 40 mesh sieves, adjust cotton dregs water content 12～18% left and right;
(2) enzymolysis: adjust cotton dregs pH4.0～5.0, add cellulase, zytase, enzyme concentration (mass ratio) is 2.5%, 1.5%, mixes, enzymolysis 16～24h, and hydrolysis temperature is controlled at 40～50 ℃;
(3) preparation solid medium: add 4%(with respect to cotton dregs mass ratio in the cotton dregs after enzymolysis, molasses, 2.5% corn steep liquor, 1.8% urea, 1.2% ammonium sulfate, 0.15% potassium dihydrogen phosphate down together), adjust moisture content in medium 30～40%, adjusting initial pH is 7.0 left and right;
(4) aspergillus activation: get fresh wheat bran, with 60 mesh sieves, sieve and remove fine powder, in wheat bran: water=1:1.2 ratio adds water, mix thoroughly to without dry powder again without clustering phenomena.After mixing thoroughly, be distributed in 2000mL triangular flask, every about 100g of the bottled wet wheat bran of 2000mL triangle, sterilizing 40～60min under 0.1MPa gauge pressure, be cooled to 35 ℃, in each triangular flask, access inclined-plane aspergillus oryzae cv-8, aspergillus niger sp-1 spore that 1-2 ring has activated, the temperature control of culture medium product is in (35 ± 1) ℃. every 12-24h shaking flask once, after growing, spore stops preparing spore suspension;
(5) solid fermentations: aspergillus oryzae, aspergillus niger are inoculated in mix in 2.5%, 4% ratio, mix 35～40 ℃ of cultivations thoroughly, aerlbic culture 3～5 days;
(6) candida utili, saccharomyces cerevisiae activation: candida utili, saccharomyces cerevisiae are received respectively in YPD culture medium, and 30 ℃, under 180r/min rotating speed, constant temperature shaking flask is 24 hours; Then with 1:100 ratio, seed liquor is received in YPD culture medium, obtained saccharomyces cerevisiae and candida utili liquid seeds;
(7) bacillus subtilis activation: adopt LB culture medium to cultivate 24h under 28～32 ℃ of conditions, make liquid seeds;
(8) secondary solid fermentation: candida utili, saccharomyces cerevisiae, bacillus subtilis are seeded to culture medium by 2.5%, 1.5%, 2.5% inoculative proportion, cultivate 2～3 days for about 28 ℃;
(9) three solid fermentations: solid lactic acid bacterium powder is seeded to culture medium by 1.5% inoculative proportion, and 26～30 ℃ of anaerobism are cultivated 24h left and right;
(10) dry: after fermentation ends, low-temperature aeration is dry;
(11) pulverize: dried fermentation cotton dregs are pulverized and got product.
Above embodiment is only for technical scheme of the present invention is described, but not is limited; Although with reference to previous embodiment, to being had been described in detail by invention, for the person of ordinary skill of the art, the technical scheme that still can record previous embodiment is modified, or part technical characterictic is wherein equal to replacement; And to these modifications or replacement, do not make the spirit and scope of essence disengaging the present invention technical scheme required for protection of appropriate technical solution.
Claims (2)
1. microorganism fermentation improves a method for cotton dregs Forage Nutritive Value, it is characterized in that specifically comprising the following steps:
(1) pretreatment: choose without the cotton dregs that go mouldy, pulverized 40 mesh sieves, adjust cotton dregs water content 12～18% left and right;
(2) enzymolysis: adjust cotton dregs pH4.0～5.0, add cellulase, zytase, enzyme concentration (mass ratio) is 2～3%, 1～2%, mixes, enzymolysis 16～24h, and hydrolysis temperature is controlled at 40～50 ℃;
(3) preparation solid medium: add 3～5%(with respect to cotton dregs mass ratio in the cotton dregs after enzymolysis, molasses, 2～3% corn steep liquors, 1.5～2% urea, 1～1.5% ammonium sulfate, 0.1～0.2% potassium dihydrogen phosphate down together), adjust moisture content in medium 30～40%, adjusting initial pH is 7.0 left and right;
(4) aspergillus activation: get fresh wheat bran, with 60 mesh sieves, sieve and remove fine powder, in wheat bran: water=1:1.0～1.3 ratio adds water, mix thoroughly to without dry powder again without clustering phenomena; After mixing thoroughly, be distributed in 2000mL triangular flask, every about 100g of the bottled wet wheat bran of 2000mL triangle, sterilizing 40～60min under 0.1MPa gauge pressure, be cooled to 35 ℃, in each triangular flask, access inclined-plane aspergillus oryzae cv-8, aspergillus niger sp-1 spore that 1-2 ring has activated, the temperature control of culture medium product is in (35 ± 1) ℃. every 12-24h shaking flask once, after growing, spore stops preparing spore suspension;
(5) solid fermentations: aspergillus oryzae, aspergillus niger are inoculated in mix in 2～3%, 3～5% ratio, mix 35～40 ℃ of cultivations thoroughly, aerlbic culture 3～5 days;
(6) candida utili, saccharomyces cerevisiae activation: candida utili, saccharomyces cerevisiae are received respectively in YPD culture medium, and 30 ℃, under 180r/min rotating speed, constant temperature shaking flask is 24 hours; Then with 1:100 ratio, seed liquor is received in YPD culture medium, obtained saccharomyces cerevisiae and candida utili liquid seeds;
(7) bacillus subtilis activation: adopt LB culture medium to cultivate 24h under 28～32 ℃ of conditions, make liquid seeds;
(8) secondary solid fermentation: candida utili, saccharomyces cerevisiae, bacillus subtilis are seeded to culture medium by 2～3%, 1～2%, 2～3% inoculative proportions, cultivate 2～3 days for about 28 ℃;
(9) three solid fermentations: solid lactic acid bacterium powder is seeded to culture medium by 1～2% inoculative proportion, and 26～30 ℃ of anaerobism are cultivated 24h left and right;
(10) dry: after fermentation ends, low-temperature aeration is dry;
(11) pulverize: dried fermentation cotton dregs are pulverized and got product.
2. microorganism fermentation according to claim 1 improves the method for cotton dregs Forage Nutritive Value, it is characterized in that in described step (3) that solid medium is without sterilizing.
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